Partial Large Subunit Ribosomal DNA Research Articles

SUPPLEMENTAL DESCRIPTION OF CABALLEROTREMA ANNULATUM (DIESING, 1850) OSTROWSKI DE NÚÑEZ AND SATTMANN, 2002 (DIGENEA: CABALLEROTREMATIDAE) FROM A NEW HOST (ELECTROPHORUS CF. VARII) AND LOCALITY (AMAZON RIVER, COLOMBIA) WITH PHYLOGENETIC ANALYSIS AND EMENDED GENERIC DIAGNOSIS.

Herein, we provide a supplemental description of Caballerotrema annulatum (Diesing, 1850) Ostrowski de Núñez and Sattmann, 2002 (Digenea: Caballerotrematidae Tkach, Kudlai, and Kostadinova, 2016) based on specimens collected from the intestine of an electric eel, Electrophorus cf. varii (Gymnotiformes: Gymnotidae) captured in the Amazon River (Colombia). This caballerotrematid can be differentiated from its congeners by the following combination of morphological features: body surface spines forming contiguous transverse rows, concentric (wrapping dorso-ventrally around body), distributing into posterior body half (vs. restricted to anterior body half in Caballerotrema brasiliensePrudhoe, 1960; indeterminate for Caballerotrema aruanenseThatcher, 1980 and Caballerotrema piscicola [Stunkard, 1960] Kostadinova and Gibson, 2001); head collar lacking projections (vs. having them in C. brasiliense, C. aruanense, and C. piscicola), narrow (head collar more narrow than maximum body width vs. the head collar being obviously wider than the body in C. brasiliense, C. aruanense, and C. piscicola); corner spines clustered (vs. corner spines distributing as 2 separated pairs in C. brasiliense, C. aruanense, and C. piscicola); pharynx approximately at level of the corner spines (vs. pharynx far anterior to corner spines in C. brasiliense, C. aruanense, and C. piscicola); and testes ovoid and nonoverlapping (C. aruanense; vs. sinuous and overlapping in C. brasiliense and C. piscicola). Based on our results, we revise the diagnosis of CaballerotremaPrudhoe, 1960 to include features associated with the shape and distribution of body surface spines, orientation and position of head collar spines, cirrus sac, seminal vesicle, oviduct, Laurer's canal, oötype, vitellarium, and transverse vitelline ducts. We performed Bayesian inference analyses using the partial large subunit ribosomal (28S) DNA gene. Our 28S sequence of C. annulatum was recovered sister to that of Caballerotrema sp. (which is the only other caballerotrematid sequence available in GenBank) from an arapaima, Arapaima gigas (Schinz, 1822) (Osteoglossiformes: Arapaimidae) in the Peruvian Amazon. Our sequence of C. annulatum comprises the only caballerotrematid sequenced tethered to a morphological description and a voucher specimen in a lending museum. The present study is a new host record and new locality record for C. annulatum. The phylogeny comprises the most resolved and taxon-rich evolutionary hypothesis for Echinostomatoidea published to date.

First report of Puccinia gibberosa on blue oatgrass (Helictotrichon sempervirens) in Canada, with taxonomic revision of the rust species

Helictotrichon sempervirens (blue oatgrass) is a perennial grass widely grown for ornamental use in Canada. Rusted blue oatgrass plants with diseased leaves covered with uredinia were collected from two locations at metropolitan Vancouver, British Columbia (BC), in summer 2022. The rust fungus was identified as Puccinia gibberosa (= P. coronata var. gibberosa) based on combined results of morphological characteristics and molecular analyses. The internal transcribed spacer two (ITS2) and partial large subunit ribosomal DNA were amplified and sequenced. The sequence obtained for the rust fungus showed 99.9% identity with the previously reported P. coronata var. gibberosa from USA. This is the first report of rust caused by P. gibberosa on blue oatgrass in Canada. Evidence from phylogenetic analyses supports the recognition of this rust at the species level. A revised description for P. gibberosa is provided, including synonyms, host range and distribution across the globe, and incorporating comprehensive molecular and morphological results for several North American and European specimens. In addition, a lectotype and an epitype are designated here for the rust species.

Metabarcoding revealed a high diversity of Amphidomataceae (Dinophyceae) and the seasonal distribution of their toxigenic species in the Taiwan Strait

The dinophyte family Amphidomataceae includes the genera Azadinium and Amphidoma. Four of these species are known to produce azaspiracids, which are lipophilic phycotoxins accumulating in shellfish. The diversity and biogeography of Amphidomataceae is far from yet resolved. Here we performed a time series sampling of both water and sediments in the Taiwan Strait from Nov. 2018 to April 2021. Metabarcoding was performed to unveil the diversity of Amphidomataceae targeting internal transcribed spacer (ITS1) region and partial large subunit ribosomal DNA (LSU rDNA D1–D3), followed by quantitative PCR (qPCR) with modified primers for Az. poporum ribotypes. The diversity of Amphidomataceae was revealed from the water samples with the aid of ITS1 and LSU based molecular phylogeny. The LSU based approach detected only a few species. In contrast, ITS1 based dataset showed eight new Azadinium clades and several ZOTUs (zero-radius operational taxonomic units) grouping together with Am. languida. Moreover, eleven known Azadinium species including three ribotypes of Az. poporum and Az. dexteroporum, and two ribotypes of Az. spinosum, were detected. The latter two species have not been reported in China before. Among these toxigenic species, Az. poporum was relevantly abundant whereas others were rare. The maximum of 209 cells L −1 of Az. poporum ribotype A was estimated using qPCR nearby Quanzhou in Nov. 2018 and 172 cells L 1 of Az. poporum ribotype B was detected far off coast in Apr. 2021. Metabarcoding on sediment samples revealed Az. poporum ribotypes B and C, but strains obtained with sediment incubation experiments yielded only ribotype B. Using qPCR about 0.2 cysts g −1 of Az. poporum ribotype B were quantified in May 2019 but cysts of Az. poporum ribotype C were not detected. Our results suggest that metabarcoding targeting ITS1 region is powerful to uncover the diversity of harmful dinophytes. Our results also highlight the rich diversity of Amphidomataceae and risk potential of azaspiracids in the Taiwan Strait and surrounding waters.

Morphologic and phylogenic characterization of two bloom-forming planktonic Prorocentrum (Dinophyceae) species and their potential distribution in the China Sea

The genus Prorocentrum includes many species that are responsible for harmful algal blooms globally, which have serious economic impacts on marine aquaculture, fisheries, and tourism, as well as major environmental and human health impacts. In this study, two bloom-forming planktonic Prorocentrum species, P. cordatum and P. sigmoides, were isolated from the coastal waters of China. Their morphology was examined in detail using light microscopy (LM) and scanning electron microscopy (SEM). The features of their periflagellar area platelet pattern, important taxonomic criteria, were revealed, and the distinctions in their morphology were compared with other closely related species. Both partial large subunit ribosomal DNA (LSU rDNA) and internal transcribed spacer region (ITS1-5.8S-ITS2) sequence-based phylogeny revealed three clades within planktonic Prorocentrum, referred to as P. micans-like (where the P. sigmoides included), P. cordatum-like, and P. triestinum-like groups. These groups showed consistencies in their thecal plate ornamentation and periflagellar area platelet pattern characteristics. Suitable habitats of P. cordatum and P. sigmoides along the coast of China were predicted with combined maximum entropy (MaxEnt) and random forest (RF) models. Highly or very highly suitable P. cordatum habitats were mainly located in the coastal waters of central to northern China. The habitat suitability range of P. sigmoides was narrower than that of P. cordatum, and concentrated near the Yangtze River Estuary. The main environmental drivers were identified by the variable importance evaluation procedure in BIOMOD 2. Mean sea surface temperature (SST) was the main distribution driver of both P. cordatum and P. sigmoides, and mean sea surface salinity (SSS), distance from coastline, and phosphorus concentrations also influenced their distribution. Ocean warming, eutrophication, and re-oligotrophication could potentially expand the distribution range of bloom-forming species. These results showed that machine learning models can play a useful role in the study of the potential distribution of harmful algal blooms.

Update of the Planktonic Diatom Genus Pseudo-nitzschia in Aotearoa New Zealand Coastal Waters: Genetic Diversity and Toxin Production.

Domoic acid (DA) is produced by almost half of the species belonging to the diatom genus Pseudo-nitzschia and causes amnesic shellfish poisoning (ASP). It is, therefore, important to investigate the diversity and toxin production of Pseudo-nitzschia species for ASP risk assessments. Between 2018 and 2020, seawater samples were collected from various sites around Aotearoa New Zealand, and 130 clonal isolates of Pseudo-nitzschia were established. Molecular phylogenetic analysis of partial large subunit ribosomal DNA and/or internal transcribed spacer regions revealed that the isolates were divided into 14 species (Pseudo-nitzschia americana, Pseudo-nitzschia arenysensis, Pseudo-nitzschia australis, Pseudo-nitzschia calliantha, Pseudo-nitzschia cuspidata, Pseudo-nitzschia delicatissima, Pseudo-nitzschia fraudulenta, Pseudo-nitzschia galaxiae, Pseudo-nitzschia hasleana, Pseudo-nitzschia multiseries, Pseudo-nitzschia multistriata, Pseudo-nitzschia plurisecta, Pseudo-nitzschia pungens, and Pseudo-nitzschia cf. subpacifica). The P. delicatissima and P. hasleana strains were further divided into two clades/subclades (I and II). Liquid chromatography-tandem mass spectrometry was used to assess the production of DA and DA isomers by 73 representative strains. The analyses revealed that two (P. australis and P. multiseries) of the 14 species produced DA as a primary analogue, along with several DA isomers. This study is the first geographical distribution record of P. arenysensis, P. cuspidata, P. galaxiae, and P. hasleana in New Zealand coastal waters.

POSTHOVITELLINUM PSILOTERMINAE N. GEN., N. SP. (DIGENEA: LISSORCHIIDAE) INFECTING THE INTESTINE OF CYCLOCHEILOS ENOPLOS (CYPRINIFORMES: CYPRINIDAE) IN THE MEKONG RIVER, VIETNAM.

Herein we describe a new species and propose a new genus, Posthovitellinum psiloterminae n. gen., n. sp. (Lissorchiidae: Asymphylodorinae), based on specimens that infect the intestine of Cyclocheilos enoplos (Bleeker, 1849) (Cypriniformes: Cyprinidae), a migratory riverine carp from the Mekong River (Dong Thap province, Vietnam). The new species is assigned to Lissorchiidae by having a combination of features: spinous tegument, subterminal oral sucker, pre-equatorial ventral sucker, median and pretesticular ovary, submarginal genital pore at level of the ventral sucker, follicular vitellarium distributing in 2 lateral fields, and lacking eyespot pigment in the adult. It cannot be assigned to any existing asymphylodorine genus because it has the combination of a well-developed cirrus-sac, an unarmed ejaculatory duct and metraterm, a follicular vitellarium distributing in 2 lateral fields located between the posterior margin of the ventral sucker and the mid-level of the testis, and a sinistral, submarginal genital pore. The new species has an elongate, claviform cirrus-sac, a single, large, elongate-oval testis at the posterior extremity of the body, operculate eggs, and an I-shaped excretory bladder with secondary branches at the level of the testis and extending anteriad to the level of the pharynx. Bayesian inference analysis of the partial large subunit ribosomal DNA gene (28S rDNA) recovered the new species sister to Asaccotrema vietnamienseSokolov and Gordeev, 2019; these species differed by 118 nucleotides (12%; 983 bp fragment). This is the first lissorchiid reported from the Mekong River; only the second from southern Vietnam; and the fourth reported from a cyprinid fish in Vietnam. The aforementioned phylogenetic analysis included previously unpublished sequences representing lissorchiids infecting the intestine of North American suckers (Cypriniformes: Catostomidae): Lissorchis cf. nelsoni from spotted sucker; Minytrema melanops (Rafinesque, 1820) and Lissorchis cf. gullaris (immature) from smallmouth buffalo, Ictiobus bubalus (Rafinesque, 1818). Asymphylodora atherinopsidisAnnereaux, 1947, herein is treated as a species incertae sedis. The 28S tree topology suggests that Lissorchiinae may comprise more than 1 lineage, but additional species are needed to confidently assert this.

Laimaphelenchus africanus n. sp. (Tylenchomorpha: Aphelenchoididae) from South Africa, a morphological and molecular phylogenetic study, with an update to the diagnostics of the genus.

A newly recovered population of the genus Laimaphelenchus from a dead maritime pine wood sample in Potchefstroom, South Africa, representing a new species, named L. africanus n. sp., is herein described and illustrated based on morphological and molecular data. The new species is mainly characterized by the following: 750–987 µm long females; a cephalic region with no disc and six cephalic lobs not divided by ribs; a 10.0–12.5 µm long stylet; four incisures in the lateral field; secretory-excretory pore (SE-pore) at slightly posterior to the nerve ring; vulva with a well-developed anterior flap, vagina with two well-developed sclerotized pieces; post-vulval uterine sac (PUS) 63–125 µm long; tail conical, 30–44 µm long, ventrally curved with a subventral stalk in terminus, lacking tubercles, with six to nine small projections at the tip in scanning electron microscopy (SEM); and rare males with 17 μm long spicules. The new species was morphologically compared to those species of the genus with a stalk in tail terminus, lacking tubercles, a vulval flap and four incisures in the lateral field viz., L. liaoningensis, L. preissii, L. simlaensis, L. sinensis, L. spiflatus, and L. unituberculus. Phylogenetically, the new species was placed into the major Laimaphelenchus clade using partial large subunit ribosomal DNA (LSU rDNA D2-D3) sequences. An overall literature review corroborated the presence of the stalk (currently with two main groups) at the tail end is the main characteristic trait delimiting the genus. A compendium based on the characters of the stalk, presence/absence of a vulval flap in females and number of the lateral lines was also established.

Morphology, ultrastructure and molecular phylogeny of cyst-producing Caladoa arcachonensis gen. et sp. nov. (Peridiniales, Dinophyceae) from France and Indonesia

ABSTRACTThe dinoflagellate order Peridiniales encompasses several well circumscribed families. However, the family level of some genera, such as Bysmatrum and Vulcanodinium, has remained elusive for many years. Four Peridinium-like strains were established from the Atlantic coast of France and North Sulawesi, Indonesia through cyst germination or isolation of single cells. The cyst-theca relationship was established on specimens from the French Atlantic. Their morphologies were examined using light, scanning and transmission electron microscopy. The cells were characterized by a much larger epitheca relative to the hypotheca, a large anterior sulcal (Sa) plate deeply intruding the epitheca and a small first anterior intercalary plate. The plate formula was identified as Po, cp, X, 4′, 3a, 7′′, 6C, 5S, 5′′′, 2′′′′, shared by Apocalathium, Chimonodinium, Fusiperidinium and Scrippsiella of the family Thoracosphaeraceae but the configuration of Sa plate and anterior intercalary plates is different. Transmission electron microscopy showed that the eyespot was located within a chloroplast comprising two rows of lipid globules and thus belongs to type A. All four strains were classified within a new genus Caladoa as C. arcachonensis gen. et sp. nov. Small subunit ribosomal DNA (SSU rDNA), partial large subunit ribosomal DNA (LSU rDNA) and internal transcribed spacer ribosomal DNA (ITS rDNA) sequences were obtained from all strains. Genetic distance based on ITS rDNA sequences between French and Indonesian strains reached 0.17, suggesting cryptic speciation in C. arcachonensis. The maximum likelihood and Bayesian inference analysis based on concatenated data from SSU and LSU rDNA sequences revealed that Caladoa is monophyletic and closest to Bysmatrum. Our results supported that Caladoa and Bysmatrum are members of the order Peridiniales but their family level remains to be determined. Our results also support that Vulcanodinium is closest to the family Peridiniaceae.

A taxonomical study of benthic Prorocentrum species (Prorocentrales, Dinophyceae) from Anse Dufour (Martinique Island, eastern Caribbean Sea)

About 30 benthic Prorocentrum species have been described, some of which producing okadaic acid and derivatives involved in diarrhetic shellfish poisoning. The western Caribbean has been extensively studied for benthic dinoflagellates associated with ciguatera, and fifteen Prorocentrum species were described from mangroves and coral reefs of Belize. In contrast, no study reported the diversity of this genus in the Eastern Caribbean, especially in the Lesser Antilles. This study adds to the biodiversity knowledge in Martinique Island by investigating one site of the Caribbean coast from 2010 to 2017. Sediment samples were collected each year in March and studied taxonomically. Identification was realized morphologically by scanning electron microscopy, while the partial large subunit (LSU) ribosomal DNA was sequenced for 42 isolated specimens (single-cells) and one strain in culture. A molecular phylogenetic analysis revealed 11 OTUs from Martinique, identified morphologically as P. concavum, P. cf. foraminosum, P. cf. tropicale, P. lima, P. hoffmannianum, P. cf. norrisianum, P. glenanicum, P. panamense, P. cf. sculptile, P. cf. fukuyoi, and P. rhathymum. Two morphospecies were also identified (P. cf. maculosum and P. cf. ruetzlerianum) but with no sequence obtained. Some species like P. cf. tropicale and P. cf. norrisinamum are sequenced for the first time. Our analysis reveals probable former misidentifications of P. cf. foraminosum and P. cf. sculptile since the sequences from Martinique form new clades and their geographical origin are closer from the type locality than any other previous studies. Further studies and sequences from the type localities are yet required to assess identifications.

Morpho-molecular diversity and phylogeny of Bysmatrum (Dinophyceae) from the South China Sea and France

ABSTRACTThe dinoflagellate genus Bysmatrum encompasses five epibenthic or tide-pool species and has been characterized by separated anterior intercalary plates. In the present study, we obtained six strains of Bysmatrum from the South China Sea and French Atlantic coast by isolating single cells/cysts from plankton and sediment samples. All strains were examined with light microscopy and scanning electron microscopy. Based on morphological observations, three strains were identified as Bysmatrum subsalsum, characterized by the elongated and rectangular first and a hexagonal second anterior intercalary plate. They differ from each other in the number of sulcal lists and the configuration of the first anterior intercalary plate. One strain was identified as Bysmatrum gregarium and the other two as Bysmatrum granulosum. The cyst-theca relationship of B. subsalsum from the French Atlantic was established by incubation of the cyst, and the geochemical composition of the cyst wall was measured through micro-Fourier transform infrared spectroscopy. Bysmatrum subsalsum from Malaysia shows a bright red stigma in the sulcal area under light microscopy, which was confirmed with transmission electron microscopy: it was identified as a type B eyespot. Small subunit ribosomal DNA (SSU rDNA), partial large subunit ribosomal DNA (LSU rDNA) and internal transcribed spacer (ITS) sequences were obtained from all six strains. The maximum likelihood and Bayesian inference analysis based on concatenated SSU, ITS and LSU sequences revealed that Bysmatrum is monophyletic and nested within Peridiniales. Our strains of B. subsalsum form a new ribotype in the molecular phylogeny (designated as ribotype B). The genetic distance based on ITS sequences among Bysmatrum species ranged from 0.34 to 0.47 and those genetic distances at the intraspecific level of B. subsalsum could reach 0.41, supporting the possibility of hidden crypticity within B. subsalsum.

Morphology, molecular phylogeny and okadaic acid production of epibenthic Prorocentrum (Dinophyceae) species from the northern South China Sea

Around 30 epibenthic Prorocentrum species have been described, but information about their biogeography is limited. Some species are able to produce okadaic acid (OA) and its derivatives, which are responsible for diarrheic shellfish poisoning (DSP). In the present study, we examined the diversity of epibenthic Prorocentrum in the northern South China Sea by isolating single cells from sand, coral, and macroalgal samples collected from 2012 to 2015. Their morphology was examined using light microscopy and scanning electron microscopy. Among 47 Prorocentrum strains, seven morphospecies were identified as P. lima, P. rhathymum, P. concavum, P. cf. emarginatum, P. fukuyoi, P. cf. maculosum and P. panamense. The latter five species have not been previously reported in Chinese waters, and this is the first record of P. panamense outside its type locality. Partial large subunit (LSU) ribosomal DNA and internal transcribed spacer region sequences were obtained and molecular phylogenetic analysis was carried out using maximum likelihood and Bayesian inference. Chinese P. cf. maculosum strains share 99.5% similarity of LSU sequences with the strain from Cuba (close to the type locality), but Chinese P. lima strains share only 96.7% similarity of LSU sequences with the strain from the type locality. P. cf. emarginatum differs from P. fukuyoi mainly in the presence/absence of marginal pores and they form a well-resolved clade together with P. sculptile. OA was detected in all Chinese strains of P. lima and P. cf. maculosum based on liquid chromatography-mass spectrometry analysis, but dinophysistoxin was produced only by two P. lima strains. Chinese strains of P. concavum, P. rhathymum, and P. panamense do not produce detectable level of OA. Our results support the wide distribution of epibenthic Prorocentrum species and highlight the potential risk of DSP in the northern South China Sea.

Cyst-motile stage relationship, morphology, ultrastructure, and molecular phylogeny of the gymnodinioid dinoflagellateBarrufeta resplendenscomb. nov., formerly known asGyrodinium resplendens, isolated from the Gulf of Mexico

In the present study, we redescribed Gyrodinium resplendens through incubation of process bearing cysts extracted from sediment collected in the northern Gulf of Mexico. The morphology and ultrastructure of the motile stage and cyst stage were examined using light microscopy, scanning electron microscopy, and transmission electron microscopy and this revealed that the species should be transferred to the genus Barrufeta. This genus differs from other gymnodinioid genera in possessing a Smurf-cap apical structure complex (ASC) and currently encompasses only one species, Barrufeta bravensis. B.resplendens shows a Smurf-cap ASC that consists of three rows of elongated vesicles with small knobs in the middle one. B.resplendens is very similar to B.bravensis in cell morphology, but can be separated using the ultrastructure such as the shape and location of nucleus and pyrenoids, which highlights the importance of ultrastructure at inter-specific level in the genus Barrufeta. The unique cysts of B.resplendens are brown and process bearing, and have a tremic archeopyle with a zigzag margin on the dorsal side of the epicyst, and not polar as in cysts of Polykrikos. The cysts do not survive the palynological treatment used here and probably have a wide distribution. Maximum-likelihood and Bayesian inference were carried out based on partial large subunit ribosomal DNA (LSU rDNA) sequences. Molecular phylogeny supports that the genus Barrufeta is monophyletic, and that the genus Gymnodinium is polyphyletic. Our results suggest that details of the ASC together with ultrastructure are potential features to subdivide the genus Gymnodinium.

Brigantedinium majusculum is the cyst of Protoperidinium sinuosum (Protoperidiniaceae, Dinophyceae)

:Two morphologically distinct species, Brigantedinium majusculum and Trinovantedinium applanatum, have been suggested as a cyst stage of Protoperidinium pentagonum; however, the cyst–theca relationships are still virtually undefined. In this study, we re-examined the motile stage of B. majusculum via thecal plate analysis together with a molecular phylogenetic analysis based on partial large-subunit ribosomal DNA gene sequences. The morphological features of motile cells germinated from B. majusculum were identical to those of Pr. sinuosum, and the molecular phylogenetic analysis showed that the germinated cells and B. majusculum are closely related to Pr. pentagonum and Pr. latissimum as a sister group. Additionally, high sequence divergences were observed between the germinated cells and B. majusculum as well as Pr. pentagonum and Pr. latissimum (18.0–19.5% according to P-values and 20.7–22.9% according to Kimura two-parameter values). On the basis of these results, our study indicates that B. majusculum should be accepted as the cyst of Pr. sinuosum.

Cyst‐theca relationships and phylogenetic positions of Peridiniales (Dinophyceae) with two anterior intercalary plates, with description of Archaeperidinium bailongense sp. nov. and Protoperidinium fuzhouense sp. nov

SummaryProtoperidinium species with two anterior intercalary plates were originally classified by Jörgensen in the subgenus Archaeperidinium and assigned to the sections Excentrica, Avellana and Archaeperidinium by Taylor, on the basis of the relative size of anterior intercalary plates and the extent of cingulum displacement. Phylogenetic relationships among these three sections have not been fully explored. Recently, Archaeperidinium was reinstated as a genus, but several species fitting the emended description have not been transferred formally as molecular data were not available. In the present study we examined the cyst‐theca relationship of seven species with two anterior intercalary plates: four species assigned to Protoperidinium and three to Archaeperidinium, from the China Sea. Partial large subunit ribosomal DNA sequences were obtained from these seven species by single‐cell polymerase chain reaction, and for the cyst of Protoperidinium stellatum from France. Two new species, Archaeperidinium bailongense and Protoperidinium fuzhouense, were described based on both theca and cyst morphology, and the cyst‐theca relationships of A. constrictum and P. abei var. rotunda were established for the first time. Maximum‐likelihood and Bayesian inference analyses revealed that P. fuzhouense was nested within the Protoperidinium sensu stricto clade despite having only six postcingular plates, and sections Excentrica, Avellana and the genus Archaeperidinium were all monophyletic. Our results suggest that the presence/absence of a sulcal fin and antapical horns and the displacement of the cingulum are the most stable characteristics of the motile stages within the respective clades.

Cyst–theca relationship and phylogenetic positions of Protoperidinium (Peridiniales, Dinophyceae) species of the sections Conica and Tabulata, with description of Protoperidinium shanghaiense sp. nov.

:The genus Protoperidinium has been divided into several sections based on the shape of the first apical plate, the number/shape of anterior intercalary plates and the presence/absence of antapical horns/spines; however, phylogenetic relationships among these sections have not been fully explored. We examined the cyst–theca relationships of seven Protoperidinium species isolated from sediments along the Chinese coast. Ten partial nuclear-encoded large subunit ribosomal DNA (LSU rDNA) sequences were obtained by single-cell polymerase chain reaction (PCR) for seven species belonging to the Conica and Tabulata sections of the genus. New cyst–theca relationships were established for Protoperidinium biconicum and Protoperidinium humile, and the former was restricted to warmer waters. Brigantedinium majusculum was identified as the cyst of Protoperidinium pentagonum; whereas, Trinovantedinium applanatum corresponded to Protoperidinium shanghaiense sp. nov. Phylogenetic analyses based on LSU rDNA sequences were carried out using maximum likelihood and Bayesian inference and revealed that the theca-based section Conica was polyphyletic, and Tabulata was monophyletic. Surprisingly, some cyst-based genera (e.g. Brigantedinium, Selenopemphix) proved to be polyphyletic.

A new holomorphic species of Mariannaea and epitypification of M. samuelsii

A new holomorphic species, Mariannaea dimorpha, is described and illustrated. The verticillate conidiophores, phialidic conidiogenous cells and aseptate microconidia that form imbricate chains indicate that it belongs to the genus Mariannaea. Sequence analyses of the combined internal transcribed spacer (ITS) and partial large subunit ribosomal DNA (28S rDNA) confirmed its taxonomic position and revealed its distinction from any morphologically similar species. In addition, the teleomorph of M. samuelsii is reported for the first time and its epitype is designated.

Taxonomy and phylogeny of the genus Cryptomonas (Cryptophyceae, Cryptophyta) from Korea

The genus Cryptomonas is easily recognized by having two flagella, green brownish color, and a swaying behavior. They have relatively simple morphology, and limited diagnostic characters, which present a major difficulty in differentiating between species of the genus. To understand species delineation and phylogenetic relationships among Cryptomonas species, the nuclear-encoded internal transcribed spacer 2 (ITS2), partial large subunit (LSU) and small subunit ribosomal DNA (rDNA), and chloroplast-encoded psbA and LSU rDNA sequences were determined and used for phylogenetic analyses, using Bayesian and maximum likelihood methods. In addition, nuclear-encoded ITS2 sequences were predicted to secondary structures, and were used to determine nine species and four unidentified species from 47 strains. Sequences of helix I, II, and IIIb in ITS2 secondary structure were very useful for the identification of Cryptomonas species. However, the helix IV was the most variable region across species in alignment. The phylogenetic tree showed that fourteen species were monophyletic. However, some strains of C. obovata had chloroplasts with pyrenoid while others were without pyrenoid, which used as a key character in few species. Therefore, classification systems depending solely on morphological characters are inadequate, and require the use of molecular data.

New dinoflagellate species Protoperidinium haizhouense sp. nov. (Peridiniales, Dinophyceae), its cyst‐theca relationship and phylogenetic position within the Monovela group

SummaryThe number of cingular plates has been used to differentiate Protoperidinium from Peridinium and related genera. Protoperidinium is characterized by the presence of three cingular plates plus a transitional plate (3C+t). However, many Protoperidinium species have been described that exhibit different cingular plate tabulations. How these species should be classified within the genus remains unclear. To address this question, the phylogenetic relationship of four Protoperidinium species, with three or four cingular plates and lacking a transitional plate, were examined in relationship to other Protoperidinium species. These four species were germinated from cysts deposited in surface sediments collected from the East China Sea, the Bohai Sea and the Yellow Sea. Three of the isolated species, P. tricingulatum, P. americanum and P. parthenopes, were described previously. The fourth is here described as P. haizhouense sp. nov. with the plate formula Po, X, 4′, 3a, 7′′, 3C, 6S, 5′′′, 2′′′′. Differences in the cyst stages of these four species, which can be taxonomically informative, were compared. Partial large subunit ribosomal DNA sequences were obtained by single‐cell polymerase chain reaction. Maximum‐likelihood and Bayesian inference showed that these four species, P. fukuyoi and Islandinium minutum form a monophyletic clade with maximal support. The genus as a whole, however, appeared polyphyletic. Our results suggest that the presence/absence of a transitional plate is significant in the phylogeny of Protoperidinium.

Morphology, ultrastructure and phylogeny of Takayama xiamenensis sp. nov. (Gymnodiniales, Dinophyceae) from the East China Sea

Gu H., Luo Z., Zhang X., Xu B. and Fang Q. 2013. Morphology, ultrastructure and phylogeny of Takayama xiamenensis sp. nov. (Gymnodiniales, Dinophyceae) from the East China Sea. Phycologia 52: 256–265. DOI: 10.2216/12-090.1A new species, Takayama xiamenensis (Kareniaceae, Dinophyceae), was isolated from Xiamen Harbor, East China Sea, and its morphology was examined using light microscopy, scanning electron microscopy, and transmission electron microscopy (TEM). The vegetative cells of T. xiamenensis were 20.0–27.0 μm long, 18.0–23.0 μm wide and 17.0–20.5 μm thick. The apical groove was deeply sigmoid, and the sulcus extended into the epicone as a distinctive finger-like protrusion. Epifluorescence microscopy revealed a large, cup-shaped nucleus, filling most of the epicone. TEM showed that the chloroplasts were enveloped by two membranes and that some chloroplasts possessed large, bulging pyrenoids. Partial large subunit ribosomal DNA sequences of two strains of T. xiamenensis were identical to the putative T. acrotrocha strains from Italy, suggesting these strains might be conspecific. The original description of T. acrotrocha from Australia does not include a finger-like protrusion, thus justifying the new species description.

Two anthozoans, Entacmaea quadricolor (order Actiniaria) and Alveopora japonica (order Scleractinia), host consistent genotypes of Symbiodinium spp. across geographic ranges in the northwestern Pacific Ocean

The actiniarian sea anemone, Entacmaea quadricolor, and the scleractinian coral, Alveopora japonica, host symbiotic dinoflagellates belonging to the genus Symbiodinium (Freudenthal). We studied the host–symbiont specificity of these two anthozoan hosts in the northwestern Pacific Ocean. Symbionts within the two hosts were identified using partial large subunit (LSU) ribosomal DNA (rDNA) and complete internal transcribed spacers (ITS) 1 rDNA regions. The host, E. quadricolor, was identified using the partial LSU rDNA molecular marker. Genetic analysis showed that E. quadricolor only harbors dinoflagellates belonging to subclade C1/3 of the genus Symbiodinium. Moreover, no genetic variation was detected among the symbionts of E. quadricolor within the study region (Korea and Japan), even though the two distant sites were separated by more than 1000 km, at collection depths of 1 m in shallow and 13–16 m in deep water. Whilst scleractinian corals host multiple Symbiodinium clades in tropical waters, A. japonica, sampled over a wide geographical range (800 km) within the study region, only hosts Symbiodinium sp. clade F3. The high specificity of endosymbionts in E. quadricolor and A. japonica within the northwestern Pacific Ocean could be accounted for because symbiotic dinoflagellates within the host anemones appear to be acquired maternally, and the Kuroshio Current might affect the marine biota of the northwestern Pacific. However, the consistency of the symbiotic relationships between these two anthozoan hosts and their endosymbionts could change after climate change, so this symbiotic specificity should be monitored.